Fig. 6. Eomes-MO2 reduces Eomes protein levels and expression of gsc and flh. (A) Western blot of in vitro transcription and translation of eomes-pCS2+ probed with the anti-Eomes antibody. Lane 1, protein produced in the presence of 5 ng of cyclops morpholino. Lane 2, protein produced without morpholino present. Lane 3, protein produced in the presence of Eomes-MO2. The amount of Eomes protein produced is not affected by the presence of cyclops morpholino but is dramatically reduced in the presence of Eomes-MO2. (B) Embryo (1 dpf), dorsal side up and anterior to the top, stained with anti-Eomes antibody. Nuclear staining in the brain is in green (arrowheads). (C) Embryo (1 dpf) in the same orientation as B that has been injected with Eomes-MO2 and stained with the anti-Eomes antibody. No staining is visible. (D) Sphere-stage embryo stained with the anti-Eomes antibody. Staining is visible in nuclei on the dorsal side of the embryo. (E) Sphere-stage embryo injected with Eomes-MO2 and stained with the anti-Eomes antibody. Staining is reduced compared to D, but nuclear-localized protein is visible on the dorsal side of the embryo (arrowhead). (F-I) Animal-pole view of shield-stage embryos. Bottom right corner indicates in situ probe used, top right corner indicates gene construct, if injected. (F) Uninjected embryo stained for gsc. (G) Eomes-MO2 injected embryo, expression of gsc is reduced. (H) Uninjected embryo stained for flh. (I) Eomes-MO2 injected embryo expression of flh is reduced.

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